微球包被手册 Particles Coating Procedures
SpheroTechnical Notes #1 - Particles Coating Procedures
Currently, there are several methods of attaching biological ligands to polystyrene particles. These methods include adsorption to plain polystyrene particles, covalent attachment to surface functionalized particles, and attachment of the ligand of interest to particles that are pre-coated with a binding protein such as Streptavidin, Protein A or Protein G. Presented in this Spherotech Technical Note are protocols such as adsorption, covalent coupling, and other methods used to attach ligands to polystyrene particles .
Procedures and Disscussion
The following information explains generalized protocols for the attachment of ligands to polystyrene particles. These protocols are easily optimized to meet the requirements for specific applications. The following protocols are developed by Spherotech for the convenience of SPHERO™ microparticle users. They are to be utilized only as initial conditions. Spherotech encourages the optimization of the coating conditions by changing the buffer, pH or reagents concentration.
In general, polyclonal antibodies are coated to polystyrene particles by adsorption without using any coupling agents. The binding of polyclonal antibodies to polystyrene particles is strong. However, care should be taken not to over load the antibodies to the particles. If over loading occurs, leaching of the coated antibody will happen during storage. This is due to the weak interaction between antibody molecules compared to the interaction of antibody molecules to the surface of polystyrene particles. As shown in Fig. 1., the optimal antibody to particle ratio for passive adsorption is between 62.5 μg to 125 μg/mL of 0.5% w/v (5.0 mg solid / mL) of 0.8 μm polystyrene particles1.
Particle coating using passive adsorption
Amino particles with ligands or proteins using EDC and covalent coupling
Carboxyl particles with avidin or other proteins using EDC covalent coupling
Covalent coupling using a two step EDC coupling protocol
Avidin particles with biotinylated proteins using affinity coupling
Covalent coupling of protein to hydroxyl particles using cyanogen bromide (CNBr)
Dimethylamino particles with DNA using ionic interaction coupling
Ligands to modified amino proteins covalent coupling using SPDP
Periodate oxidation of polysaccharide and coupling to amino particles
Carboxyl polystyrene particles with amino modified oligonucleotides